Harlow K W, Nygaard P, Hove-Jensen B
Department of Protein Chemistry, University of Copenhagen, Denmark.
J Bacteriol. 1995 Apr;177(8):2236-40. doi: 10.1128/jb.177.8.2236-2240.1995.
The Escherichia coli gsk gene encoding guanosine kinase was cloned from the Kohara gene library by complementation of the E. coli gsk-1 mutant allele. The cloned DNA fragment was sequenced and shown to encode a putative polypeptide of 433 amino acids with a molecular mass of 48,113 Da. Minicell analysis established the subunit M(r) as 43,500. Primer extension analysis indicated the presence of an adequate Pribnow box and suggested that the transcript contained a 110-base leader sequence. Strains harboring the gsk gene on multicopy plasmids overexpressed both guanosine and inosine kinase activities. N-terminal sequence and amino acid composition analyses of the 43,500-M(r) polypeptide band confirmed the correct reading frame assignment and the identity of this band as the gsk gene product. Comparison of the amino acid sequence with the protein database revealed similarity to regions of other mononucleotide-utilizing enzymes.
通过对大肠杆菌gsk - 1突变等位基因进行互补，从Kohara基因文库中克隆出编码鸟苷激酶的大肠杆菌gsk基因。对克隆的DNA片段进行测序，结果表明其编码一个推定的由433个氨基酸组成的多肽，分子量为48,113道尔顿。小细胞分析确定该亚基的相对分子质量为43,500。引物延伸分析表明存在合适的Pribnow框，并提示该转录本包含一个110个碱基的前导序列。携带多拷贝质粒上gsk基因的菌株同时过度表达鸟苷激酶和肌苷激酶活性。对相对分子质量为43,500的多肽条带进行N端序列和氨基酸组成分析，证实了正确的读框分配以及该条带作为gsk基因产物的身份。将氨基酸序列与蛋白质数据库进行比较，发现它与其他利用单核苷酸的酶的区域具有相似性。
